CD8+Tcellsareessentialcomponentsoftheimmunesystem.TheyfunctionthroughTCRrecognitionofantigens(peptideepitopes)presentedbytheclassIMHC(HLAinhumans)molecules.Thus,TCRrecognitionofcognateMHC/peptidecomplexcanbeutilizedforthedetectionofantigen-specificCD8+Tcells.DuetotheintrinsiclowaffinityofMHC/TCRinteraction,thedetectionofantigenspecificTcellsrequiresenhancedbindingAviditybymultimerizationofMHC/peptidemonomers,ensuringtheinteractionofmultipleMHC/TCRsimultaneously.ThiscanbeaccomplishedbyusingstreptavidintotetramerizebiotinylatedMHCclassIcomplexes.Intheabsenceofproperpeptidebinding,theHLAclassIcomplexwillquicklydisassociate.Flex-T™UVX(UltravioletExchange)monomerisdesignedasabiotinylatedα-chain,associatedwiththeβ2-microglobulinchain,andstABIlizedbyaUV-labilepeptide.TheC-terminusoftheα-chainissite-specificallybiotinylatedbytheenzymeBirA.Whenthecomplex(monomer)isirrADIatedwithUVinthepresenceofanewpeptidewithproperaffinity,itwillresultinanetpeptideexchange.Thenewpeptidereplacesthepre-loadedUV-labilepeptideandformsanewMHCmonomer.Thenewmonomerscanbethentetramerizedandusedforlabelingantigen-specificTcellsorotherstudies.
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